Pierce™ Agarose ChIP Kit
Pierce™ Agarose ChIP Kit
Thermo Scientific™

Pierce™ Agarose ChIP Kit

The Thermo Scientific Pierce Agarose ChIP Kit provides a complete set of reagents and a simple, fast and reproducible protocolRead more
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Catalog number 26156
Price (USD)
764.00
Each
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Price (USD)
764.00
Each
Add to cart
The Thermo Scientific Pierce Agarose ChIP Kit provides a complete set of reagents and a simple, fast and reproducible protocol to perform chromatin immunoprecipitation (ChIP) assays.

Features of the Agarose ChIP Kit:

• Simple and fast ChIP assay protocol (< 8 hours)
• Highly efficient isolation and lysis of nuclei
• Easy and reproducible enzymatic digestion
• Low-background and high-binding capacity Protein A/G agarose resin
• Highly specific positive controls included: RNA polymerase II antibody and GAPDH PCR primers
• Fast and reproducible spin-column format
• High-recovery DNA purification
• ChIP-validated antibodies available

The Pierce Agarose ChIP Kit contains sufficient reagents to perform 30 ChIP assays with appropriate controls using an optimized protocol and a convenient spin-column format. ChIP-validated and quality-guaranteed antibodies are also available for use the the Pierce Agarose ChIP Kit. Because antibodies that work for western blotting may not function in ChIP assays, we have assembled our most popular ChIP-validated monoclonal and polyclonal antibodies here to help you move from experimental setup to results more quickly.

The strength of the ChIP assay is its ability to capture a snapshot of specific protein-DNA interactions as they occur in living cells, and then quantitate the interactions using standard or quantitive PCR. A successful ChIP assays requires a number of critical steps to be carried out (crosslinking, chromatin preparation, immunoprecipitation) prior to detecting the target genomic DNA sequence. Individually, each step in the ChIP assay protocol can be time consuming to optimize. The Pierce Agarose ChIP Assay Kit simplifies the process, making accurate and reproducible results easier to obtain.

To perform ChIP assays with the Pierce Agarose ChIP Kit, protein-DNA complexes are stabilized and then extracted. In vivo crosslinking is traditionally achieved with formaldehyde. Crosslinking performed directly in cells locks in the protein-DNA complexes, trapping these unstable and sometimes transient interactions. To lyse, extract and solubilize the crosslinked complexes, the ChIP assay kit includes the Thermo Scientific Chromatin Prep Module (also sold separately, Part No. 26158). Together, this complete ChIP assay kit provides a simple, reliable and convenient means for isolating chromatin-bound DNA and enriching samples for the proteins of interest with less than 15% contamination from other cellular compartments - without a Dounce homogenizer.

Related Products
Pierce™ Chromatin Prep Module
Pierce™ ChIP-Grade Protein A/G Plus Agarose
Proteinase K
For Research Use Only. Not for use in diagnostic procedures.
Specifications
Binding Capacity>50 mg Human IgG/mL resin binding capacity
Capacity (Metric)≥50 mg Human IgG per mL Resin Binding
DescriptionPierce Agarose ChIP Kit
Product LinePierce™
Quantity30 Reaction Kit
TypeChIP Kit
Sufficient For30 Reactions
FormatKit
Unit SizeEach
Contents & Storage
Sufficient For: 30 complete chromatin IP assay experiments• ChIP Grade Protein A/G Plus Agarose, 0.65 mL (store at 4°C)
• IP Dilution/Wash Buffer (5X), 11 mL (store at 4°C)
• IP Wash Buffer 3 (5X), 4.5 mL (store at 4°C)
• IP Elution Buffer (2X), 4.5 mL (store at 4°C)
• Column Accessory Pack, 3 packs (store at room temperature or 4°C)
• Microcentrifuge Tubes (1.5mL), 75 tubes (store at room temperature or 4°C)
• DNA Clean-Up Column, 40 columns (store at room temperature or 4°C)
• DNA Column Binding Solution, 30 mL (store at room temperature or 4°C)
• DNA Column Wash Solution, 6 mL (store at room temperature or 4°C)
• pH Indicator, 0.8 mL (store at room temperature or 4°C)
• DNA Column Elution Solution, 5 mL (store at room temperature or 4°C)
• Anti-RNA Polymerase II Antibody, 25 μL (store at -20°C)
• Normal Rabbit IgG (1 mg/mL), 10 μL (store at -20°C)
• ChIP Positive Control Primers (GAPDH promoter, Human-specific), 100 μL (store at -20°C)
• Pierce Chromatin Prep Module (Part No. 26158) (store Proteinase K and Micrococcal Nuclease at -20°C. Store all other components at 4°C).

Frequently asked questions (FAQs)

I am having trouble with leaking columns while using the Pierce Agarose ChIP Kit (Cat. No. 26156). How can I resolve this issue?

It is common to experience leaking when using the columns, even when the plug is pushed in very tightly. We recommend wrapping the plug with parafilm to prevent leaking.

Find additional tips, troubleshooting help, and resources within our Protein Assays and Analysis Support Center.

With my ChIP assay (Agarose ChIP Kit/Magnetic ChIP Kit), the qPCR signal (cycle threshold) from my positive and negative controls show no difference or are very close. Why is this?

-Verify that your specific antibody (if not using the kit-provided RNA polymerase II antibody) is validated for IP. Ideally, a ChIP validated antibody is the best, but an antibody for IP has a good chance of working in ChIP.
-Ensure that your chromatin is properly digested (see Appendix A in the manual). Too much digestion as well as too little digestion will affect the success of the ChIP reaction.
-Ensure that all the chromatin has been released from the nuclei. When following the Magnetic ChIP kit instructions, MNase digestion of 4x106 cells followed by sonication to lyse the nuclei, yields about 20-50 µg for the IP. This same sequence can be used with the Agarose ChIP Kit as well. It is recommended that you start with 2–4 x 106 cells per ChIP reaction. Once a successful ChIP has been run at this cell number, it is possible to decrease the cell amount empirically. We have seen good results using as little at 10,000 cells, but this entirely depends on the cell line, target, and antibody.
-Ensure that enough DNA was used for qPCR. Typically, 30-80 ng of DNA is a good range.

With my Agarose ChIP Kit, what can cause no or low PCR signal in the experimental IP samples?

Here are possible causes and solutions:

- Insufficient amount of sample DNA added to the PCR reaction: Add more sample DNA to the PCR reaction.
- Insufficient amount of antibody added to the IP: Add more antibody to the IP.
- Antibody did not function in an IP: Verify that the antibody is qualified for CHIP or IP applications and has been handled and stored properly.

Note: Increasing the stringency of the nuclear lysis is not recommended unless there is no signal or low signal-to-noise in the IP. Excess sample can result in high background decreasing the signal of the specific signal.

With my Agarose ChIP Kit, what can cause PCR signal of the positive and negative control IP samples to be equivalent?

Here are possible causes and solutions:

- Insufficient washing of the IP complex: Include an additional wash with Buffers 2 and 3.
- Excess chromatin or antibody added to the IP: Add less chromatin or antibody.
- PCR amplification was measured outside the linear range of amplification: Decrease the number of amplification cycles used in the PCR reaction.

With my Agarose ChIP Kit, what can cause no or low PCR signal in the positive control IP samples?

Here are possible causes and solutions:

- Insufficient chromatin amount in the IP reaction: Use at least 25 µg of chromatin for each IP.
- Insufficient antibody incubation time: Incubate antibody overnight.
- Incomplete elution from the Protein A/G agarose resin: Perform elution at 65 degrees C and increase frequency of mixing.

View all Pierce™ Agarose ChIP Kit FAQs